Human complement fragment 5a ELISA kit

Attention: The products sold by our companyABCAM agent for human complement fragment 5a ELISA kit C5a reagent kitIt is only used for scientific research experiments and cannot be used for food or clinical use, edible drugs, etc.
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ELISAExperimental precautions
　　1.During the formal experiment, positive and negative controls should be used to control the experimental conditions, and two copies of the test sample should be prepared to ensure the accuracy of the experimental results. Sometimes a high background indicates non-specific reactions, and sheep serum, rabbit serum, orBSAWaiting for closure.
　　2.inELISAIt is important to choose various experimental conditions, including:
　　(1)Selection of Solid Phase Carriers: Many substances can be used as solid phase carriers, such as polyvinyl chloride, polystyrene, polyacrylamide, and cellulose. Its form can be concave hole flat plate, test tube, bead, etc. The currently commonly used ones are40Hollow polystyrene perforated plate. Regardless of the type of carrier, it can be screened before use: coat it with an equal amount of antigen, conduct the reaction under the same experimental conditions, observe whether the color reaction is homogeneous, and determine whether its adsorption performance is good based on this.
（2)Coated with antibodies(Or antigen)Choice: Apply antibodies(Or antigen)When adsorbed on the surface of a solid carrier, it is required to have good purity,General requirements for adsorptionPHin9.0～9.6between. The adsorption temperature, time, and protein content also have a certain impact,Usually used4℃18～24Hour. The appropriate concentration of protein coating needs to be titrated: using different protein concentrations(0.1、1.0and10Mg／mlwait)After coating, observe the positive specimens under the same experimental conditionsODValue. choiceODvalueThe concentration that is high but low in protein content. For most proteins, it is usually1～10Mg／ml.
　　(3)Selection of enzyme labeled antibody working concentration: First, use directELISATitration of preliminary potency using the method(See enzyme labeled antibody section)Then fix other conditions or adopt“Phalanx method”(The reference material for the package, the sample to be tested, and the enzyme labeled antibody have different dilutions)Accurately titrate its working concentration in the formal experimental system.
　　(4)The selection of substrates and hydrogen donors for enzymes: The requirements for selecting hydrogen donors are low cost, safety, clear color reaction, and colorless nature. Some hydrogen donors(as ifOPDwait)It has potential carcinogenic effects and should be protected. Conditional individuals should use non carcinogenic and highly sensitive hydrogen donors, such asTMBandABTSIt is currently a satisfactory hydrogen donor. After a period of substrate action, strong acid or strong base should be added to terminate the reaction. The usual substrate action time is10-30Minutes are appropriate. The substrate solution must be freshly prepared, especiallyH2O2Add before use.